Wendy Martens

A comparative evaluation of Cone Beam Computed Tomography (CBCT) and Multi-Slice CT (MSCT): Part I. On subjective image quality

AIMS To compare image quality and visibility of anatomical structures in the mandible between five Cone Beam Computed Tomography (CBCT) scanners and one Multi-Slice CT (MSCT) system. MATERIALS AND METHODS One dry mandible was scanned with five CBCT scanners (Accuitomo 3D, i-CAT, NewTom 3G, Galileos, Scanora 3D) and one MSCT system (Somatom Sensation 16) using 13 different scan protocols. Visibility of 11 anatomical structures and overall image noise were compared between CBCT and MSCT. Five independent observers reviewed the CBCT and the MSCT images in the three orthographic planes (axial, sagittal and coronal) and assessed image quality on a five-point scale. RESULTS Significant differences were found in the visibility of the different anatomical structures and image noise level between MSCT and CBCT and among the five CBCT systems (p=0.0001). Delicate structures such as trabecular bone and periodontal ligament were significantly less visible and more variable among the systems in comparison with other anatomical structures (p=0.0001). Visibility of relatively large structures such as mandibular canal and mental foramen was satisfactory for all devices. The Accuitomo system was superior to MSCT and all other CBCT systems in depicting anatomical structures while MSCT was superior to all other CBCT systems in terms of reduced image noise. CONCLUSIONS CBCT image quality is comparable or even superior to MSCT even though some variability exists among the different CBCT systems in depicting delicate structures. Considering the low radiation dose and high-resolution imaging, CBCT could be beneficial for dentomaxillofacial radiology.

Mesenchymal stem/stromal cells as a pharmacological and therapeutic approach to accelerate angiogenesis.

Mesenchymal stem cells or multipotent stromal cells (MSCs) have initially captured attention in the scientific world because of their differentiation potential into osteoblasts, chondroblasts and adipocytes and possible transdifferentiation into neurons, glial cells and endothelial cells. This broad plasticity was originally hypothesized as the key mechanism of their demonstrated efficacy in numerous animal models of disease as well as in clinical settings. However, there is accumulating evidence suggesting that the beneficial effects of MSCs are predominantly caused by the multitude of bioactive molecules secreted by these remarkable cells. Numerous angiogenic factors, growth factors and cytokines have been discovered in the MSC secretome, all have been demonstrated to alter endothelial cell behavior in vitro and induce angiogenesis in vivo. As a consequence, MSCs have been widely explored as a promising treatment strategy in disorders caused by insufficient angiogenesis such as chronic wounds, stroke and myocardial infarction. In this review, we will summarize into detail the angiogenic factors found in the MSC secretome and their therapeutic mode of action in pathologies caused by limited blood vessel formation. Also the application of MSC as a vehicle to deliver drugs and/or genes in (anti-)angiogenesis will be discussed. Furthermore, the literature describing MSC transdifferentiation into endothelial cells will be evaluated critically.

Angiogenic Properties of Human Dental Pulp Stem Cells

Angiogenesis, the formation of capillaries from pre-existing blood vessels, is a key process in tissue engineering. If blood supply cannot be established rapidly, there is insufficient oxygen and nutrient transport and necrosis of the implanted tissue will occur. Recent studies indicate that the human dental pulp contains precursor cells, named dental pulp stem cells (hDPSC) that show self-renewal and multilineage differentiation capacity. Since these cells can be easily isolated, cultured and cryopreserved, they represent an attractive stem cell source for tissue engineering. Until now, only little is known about the angiogenic abilities and mechanisms of the hDPSC. In this study, the angiogenic profile of both cell lysates and conditioned medium of hDPSC was determined by means of an antibody array. Numerous pro-and anti-angiogenic factors such as vascular endothelial growth factor (VEGF), monocyte chemotactic protein-1 (MCP-1), plasminogen activator inhibitor-1 (PAI-1) and endostatin were found both at the mRNA and protein level. hDPSC had no influence on the proliferation of the human microvascular endothelial cells (HMEC-1), but were able to significantly induce HMEC-1 migration in vitro. Addition of the PI3K-inhibitor LY294002 and the MEK-inhibitor U0126 to the HMEC-1 inhibited this effect, suggesting that both Akt and ERK pathways are involved in hDPSC-mediated HMEC-1 migration. Antibodies against VEGF also abolished the chemotactic actions of hDPSC. Furthermore, in the chicken chorioallantoic membrane (CAM) assay, hDPSC were able to significantly induce blood vessel formation. In conclusion, hDPSC have the ability to induce angiogenesis, meaning that this stem cell population has a great clinical potential, not only for tissue engineering but also for the treatment of chronic wounds, stroke and myocardial infarctions.

Effect of isolation methodology on stem cell properties and multilineage differentiation potential of human dental pulp stem cells

Dental pulp stem cells (DPSCs) are an attractive alternative mesenchymal stem cell (MSC) source because of their isolation simplicity compared with the more invasive methods associated with harvesting other MSC sources. However, the isolation method to be favored for obtaining DPSC cultures remains under discussion. This study compares the stem cell properties and multilineage differentiation potential of DPSCs obtained by the two most widely adapted isolation procedures. DPSCs were isolated either by enzymatic digestion of the pulp tissue (DPSC-EZ) or by the explant method (DPSC-OG), while keeping the culture media constant throughout all experiments and in both isolation methods. Assessment of the stem cell properties of DPSC-EZ and DPSC-OG showed no significant differences between the two groups with regard to proliferation rate and colony formation. Phenotype analysis indicated that DPSC-EZ and DPSC-OG were positive for CD29, CD44, CD90, CD105, CD117 and CD146 expression without any significant differences. The multilineage differentiation potential of both stem cell types was confirmed by using standard immuno(histo/cyto)chemical staining together with an in-depth ultrastructural analysis by means of transmission electron microscopy. Our results indicate that both DPSC-EZ and DPSC-OG could be successfully differentiated into adipogenic, chrondrogenic and osteogenic cell types, although the adipogenic differentiation of both stem cell populations was incomplete. The data suggest that both the enzymatic digestion and outgrowth method can be applied to obtain a suitable autologous DPSC resource for tissue replacement therapies of both bone and cartilage.

Human dental pulp stem cells can differentiate into Schwann cells and promote and guide neurite outgrowth in an aligned tissue-engineered collagen construct in vitro

In the present study, we evaluated the differentiation potential of human dental pulp stem cells (hDPSCs) toward Schwann cells, together with their functional capacity with regard to myelination and support of neurite outgrowth in vitro. Successful Schwann cell differentiation was confirmed at the morphological and ultrastructural level by transmission electron microscopy. Furthermore, compared to undifferentiated hDPSCs, immunocytochemistry and ELISA tests revealed increased glial marker expression and neurotrophic factor secretion of differentiated hDPSCs (d‐hDPSCs), which promoted survival and neurite outgrowth in 2‐dimensional dorsal root ganglia cultures. In addition, neurites were myelinated by d‐hDPSCs in a 3‐dimensional collagen type I hydrogel neural tissue construct. This engineered construct contained aligned columns of d‐hDPSCs that supported and guided neurite outgrowth. Taken together, these findings provide the first evidence that hDPSCs are able to undergo Schwann cell differentiation and support neural outgrowth in vitro, proposing them to be good candidates for cell‐based therapies as treatment for peripheral nerve injury.—Martens, W., Sanen, K., Georgiou, M., Struys, T., Bronckaers, A., Ameloot, M., Phillips, J., Lambrichts, I. Human dental pulp stem cells can differentiate into Schwann cells and promote and guide neurite outgrowth in an aligned tissue‐engineered collagen construct in vitro. FASEB J. 28, 1634–1643 (2014). www.fasebj.org

Dental stem cells and their promising role in neural regeneration: an update

IntroductionStem cell-based therapies are considered to be a promising treatment method for several clinical conditions such as Alzheimers disease, Parkinsons disease, spinal cord injury, and many others. However, the ideal stem cell type for stem cell-based therapy remains to be elucidated.DiscussionStem cells are present in a variety of tissues in the embryonic and adult human body. Both embryonic and adult stem cells have their advantages and disadvantages concerning the isolation method, ethical issues, or differentiation potential. The most described adult stem cell population is the mesenchymal stem cells due to their multi-lineage (trans)differentiation potential, high proliferative capacity, and promising therapeutic values. Recently, five different cell populations with mesenchymal stem cell characteristics were identified in dental tissues: dental pulp stem cells, stem cells from human exfoliated deciduous teeth, periodontal ligament stem cells, dental follicle precursor cells, and stem cells from apical papilla.ConclusionEach dental stem cell population possesses specific characteristics and advantages which will be summarized in this review. Furthermore, the neural characteristics of dental pulp stem cells and their potential role in (peripheral) neural regeneration will be discussed.

Macro‐ and micro‐anatomical, histological and computed tomography scan characterization of the nasopalatine canal

AIM To determine the human anatomic variability of the nasopalatine canal and determine its characteristics using an anatomical, histological and computed tomography (CT) scan evaluation. MATERIALS AND METHODS Measurements for the canal characteristics were carried out on 163 dry human skulls and 120 upper jaw spiral CT scans, taken from patients for pre-operative planning purposes of implant placement in the incisor region. Furthermore, four human cadaver specimens were imaged using a high-resolution magnetic resonance imaging (HR-MRI) unit. Afterwards, these specimens were serially sectioned for histological examination to evaluate the nasopalatine canal region and its content. RESULTS The nasopalatine canal anatomy showed a large variability in morphology and dimensions, with the canal branching in up to four canals at the level of the nose. The canal diameter was on average 3.3 mm (+/-0.9 mm SD), and typically enlarged by age and male gender (p<0.05). HR-MRI and histological sections enabled to identify the neurovascular structures within the canals. CONCLUSIONS The large anatomic variations, the increased canal dimensions with age and the neurovascular canal content are all factors favouring a thorough three-dimensional planning before surgery, such as implant placement, of the anterior maxillary region.

Neurogenic Maturation of Human Dental Pulp Stem Cells Following Neurosphere Generation Induces Morphological and Electrophysiological Characteristics of Functional Neurons

Cell-based therapies are emerging as an alternative treatment option to promote functional recovery in patients suffering from neurological disorders, which are the major cause of death and permanent disability. The present study aimed to differentiate human dental pulp stem cells (hDPSCs) toward functionally active neuronal cells in vitro. hDPSCs were subjected to a two-step protocol. First, neuronal induction was acquired through the formation of neurospheres, followed by neuronal maturation, based on cAMP and neurotrophin-3 (NT-3) signaling. At the ultrastructural level, it was shown that the intra-spheral microenvironment promoted intercellular communication. hDPSCs grew out of the neurospheres in vitro and established a neurogenic differentiated hDPSC culture (d-hDPSCs) upon cAMP and NT-3 signaling. d-hDPSCs were characterized by the increased expression of neuronal markers such as neuronal nuclei, microtubule-associated protein 2, neural cell adhesion molecule, growth-associated protein 43, synapsin I, and synaptophysin compared with nondifferentiated hDPSCs. Enzyme-linked immunosorbent assay demonstrated that the secretion of brain-derived neurotrophic factor, vascular endothelial growth factor, and nerve growth factor differed between d-hDPSCs and hDPSCs. d-hDPSCs acquired neuronal features, including multiple intercommunicating cytoplasmic extensions and increased vesicular transport, as shown by the electron microscopic observation. Patch clamp analysis demonstrated the functional activity of d-hDPSCs by the presence of tetrodotoxin- and tetraethyl ammonium-sensitive voltage-gated sodium and potassium channels, respectively. A subset of d-hDPSCs was able to fire a single action potential. The results reported in this study demonstrate that hDPSCs are capable of neuronal commitment following neurosphere formation, characterized by distinct morphological and electrophysiological properties of functional neuronal cells.

Ultrastructural and immunocytochemical analysis of multilineage differentiated human dental pulp- and umbilical cord-derived mesenchymal stem cells

Mesenchymal stem cells (MSCs) are one of the most promising stem cell types due to their availability and relatively simple requirements for in vitro expansion and genetic manipulation. Besides the well-characterized MSCs derived from bone marrow, there is growing evidence suggesting that dental pulp and the umbilical cord matrix both contain a substantial amount of cells having properties similar to those of MSCs. In order to assess the potential of dental pulp-derived MSCs (DPSC) and umbilical cord-derived MSCs (UCSC) in future clinical applications, it is essential to gain more insight into their differentiation capacity and to evaluate the tissues formed by these cells. In the present study, the morphological and ultrastructural characteristics of DPSC and UCSC induced towards osteogenic, adipogenic, and chondrogenic lineages were investigated. Cultured DPSC and UCSC showed a similar expression pattern of antigens characteristic of MSCs including CD105, CD29, CD44, CD146, and STRO-1. Under appropriate culture conditions, both DPSC and UCSC showed chondrogenic and osteogenic potential. Adipogenesis could be only partially induced in DPSC resulting in the de novo expression of fatty acid binding protein (FABP), whereas UCSC expressed FABP combined with a very high accumulation of lipid droplets in the cytoplasm. Our results demonstrate, at the biochemical and ultrastructural level, that DPSC display at least bilineage potential, whereas UCSC, which are developmentally more primitive cells, show trilineage potential. We emphasize that transmission electron microscopical analysis is useful to elucidate detailed structural information and provides indisputable evidence of differentiation. These findings highlight their potential therapeutic value for cell-based tissue engineering.

Expression Pattern of Basal Markers in Human Dental Pulp Stem Cells and Tissue

Dental pulp stem cells (DPSC) have been characterized as a multipotent stem cell population, with the ability to differentiate into mesodermal and neural cell lineages. Although ‘de novo’ expression of neural markers after differentiation is mostly considered as proof of differentiation, expression of these markers in undifferentiated DPSC is not well described. Therefore, an immunocytochemical analysis was performed to evaluate the neural marker expression of undifferentiated human DPSC (hDPSC) in in vitro cultures. Undifferentiated hDPSC uniformly expressed neural markers β-III-tubulin, S100 protein and synaptophysin. A subset of the population showed a positive immune-reactivity for galactocerebroside, neurofilament and nerve growth factor receptor p75. Furthermore, the location of possible stem cell niches, present in young dental pulp tissue, was determined by means of immunohistochemistry based on mesenchymal and neural marker expression. The results demonstrated the presence of a perivascular niche and a second stem cell niche at the cervical area. In adult dental pulp, only a perivascular niche could be observed. Based on the expression of neural markers in naïve DPSC, it has to be taken into account that not only the marker expression upon neural differentiation must be analyzed, but an ultrastructural analysis of the morphological changes and functional studies must also be performed to confirm a successful differentiation.